A reliable, simple, and cost-effective method for the immobilization of relatively short (12-30 mer) oligonucleotide probes to 96-well polystyrene plates was required in our laboratory for use in DNA hybridization-based assays. We compared three different approaches to achieve this immobilization. Two of them are modifications of previously published procedures, requiring the use of modified oligonucleotides and/or modified plates. These were compared to a method developed in our laboratory, whereby passive immobilization occurs by incubation in the presence of salt or a cationic detergent. While all methods resulted in the productive binding of the DNA probes and could therefore be used for hybridization, only the passive immobilization approach met our strict performance criteria and was implemented for use in our DNA genotyping laboratory. It was found that the immobilization in the presence of cationic detergents takes place via a unique mechanism.